Computer-designed reagent targets lysine for protein modification

At pH 8.0, a methylsulfonyl acrylate reagent modifies a protein at its most acidic lysine (black). Additional molecules (red ball) can be added by an aza-Michael ligation reaction.

Credit: J. Am. Chem. Soc.

At pH 8.0, a methylsulfonyl acrylate reagent modifies a protein at its most acidic lysine (black). Additional molecules (red ball) can be added by an aza-Michael ligation reaction.

Credit: J. Am. Chem. Soc.

Chemists modify protein sidechains to modulate the biomolecules’ functions. Researchers are usually stuck modifying cysteines because the amino acid’s sulfide side chain makes it the most reactive amino acid. But cysteines are relatively rare, representing ~1.9% of the amino acids in human proteins. Cysteines can be installed using genetic engineering, but researchers would like to modify fully functional native proteins.

An international team of researchers led by Gonçalo J. L. Bernardes of the University of Cambridge and the Institute of Molecular Medicine Lisbon and Gonzalo Jiménez-Osés of the University of Rioja has now discovered a sulfonyl acrylate reagent that should give researchers more options for modifying proteins. The reagent selectively targets only the most reactive lysine in a protein—not other lysines or buried cysteines (J. Am. Chem. Soc. 2018, DOI: 10.1021/jacs.7b12874). Lysines are nucleophilic and much more abundant than cysteines, representing ~5.9% of the amino acids in human proteins.

The terminal amine of the reactive lysine attacks the reagent, eliminating its sulfonyl group and adding a new acrylate group to the protein. As predicted by computer simulations, the reagent forms a transient hydrogen bond between its sulfonyl group and the terminal amino group of the lysine, dramatically accelerating the reaction rate. The added acrylate can be used to further modify the protein via an aza-Michael ligation with a molecule containing a reactive amine. The researchers used the reagent to modify five proteins, including a full-length antibody.

“There is a significant need to develop new chemistries for the regio- and chemoselective modification of proteins,” says Bradley L. Pentelute of Massachusetts Institute of Technology, who is also working to develop such methods. “This advance will certainly change how we think about protein modification and inspire future advancements in this area.”

The researchers are working on extending the method to attach two complementary payloads onto an antibody using orthogonal conjugation methods that target, for example, lysine and cysteine.